Introduction: Infertility is a multifactorial syndrome with various genetic and non-genetic factors. Despite methodological advancements, the possible causes for infertility are still unknown for more than 25 percent of cases (idiopathic infertility). Male factor infertility accounts for 50 percent of infertile couples. The human KISS1 GENE is located on the long arm of chromosome 1 and has a pivotal role in metastasis control. Since 2003, its importance in puberty, ovulation and fertility has been revealed. Objective: This study was conducted to investigate the relationship between codon 81 polymorphism of the gene and male idiopathic male infertility. Materials and Methods: In this molecular study, 50 patients and 36 healthy fertile men (as controls) were tested for polymorphism of pro 81 Arg. DNA was extracted from 2ml of peripheral blood samples from the patients and controls (having obtained written informed consents from them). Allele specific PCR method was applied to determine the codon polymorphism. Results: Among infertile cases, the distribution of genotypes was as follows: 5% were Pro/Pro, 84% were Arg/Pro and 5% were pro/pro. As for the controls, the distributions were: 0% Pro/Pro, 100% Arg/Pro and 0% Arg/Arg. The results and statistical analyses revealed that the genotype frequencies observed between healthy and patient groups, were significantly different between the two groups (P = 0.0418). However, difference in the distributions of alleles (Pro/Arg) in patient and control groups was not significant (P = 0.9170). Conclusion: In this study, Pro/Pro and Arg/Arg homozygote genotypes in 81 codon of KISS1 gene can be attributed to idiopathic male infertility. Overall, the role of genetic composition between the studied polymorphism and susceptibility to male idiopathic infertility may be considered as effective. However, the results may be different when selecting different genetic pools or significant changes in a population size. Thus, the conclusion was not solid and more studies are needed for functional validation of the polymorphism.